Respiratory viral infections are the third leading cause of death worldwide and are a WHO priority for vaccine development. For many years the scientific community has focused enormous efforts on the development of therapeutic and prophylactic vaccines that elicit CD8 T cell responses with the promise that such interventions will be sufficient to confer protective effects and potentially offset any deficiency in the antibody arm of the anti- viral response. However, to date the development of effective T cell vaccines against respiratory viruses remains elusive. For safety reasons, most vaccination strategies use replication-incompetent, or replication competent highly attenuated, viral vectors. However, there is considerable literature acknowledging that strongly replicating viruses are better at inducing long-term protective CD8 mediated immunity than attenuated viruses. Therefore, the effect of reduced virulence on immunogenicity becomes a central issue when attempting to derive a truly effective T cell vaccine that incorporates attenuated vectors. In this application, we will investigate the molecular basis that differentiates attenuated from non-attenuated vaccines by focusing on vaccine virus (VACV). We will use several clinically relevant natural and recombinant VACV variants that differ in their ability to modulate host immune responses and test the idea that the level of virus replication, influenced by virulence and evasion tactics, can lead to differential expression and selective use of essential activating and inhibitory receptors on CD8 T cells, to allow efficient immune responses and persistence of memory. Specifically, we will test the idea that generation of protective anti-viral CD8 T cell responses in the lung are highly regulated by the tumor-necrosis-factor receptor (TNFR) family member, herpes virus-entry mediator (HVEM). Recently it was found that HVEM could act as a molecular switch between pro inflammatory and inhibitory signaling by respectively binding with its endogenous ligand (LIGHT) from the TNF family and ITIM-containing inhibitory molecule B- and T-lymphocyte attenuator (BTLA) from the Ig like CD28/B7 family. The crosstalk between these two different families and especially between co-stimulatory and co-inhibitory receptors has raised many new questions with regards to the precise mechanisms of immune modulation through these interactions. Directly relevant to this proposal, the precise contribution of HVEM interacting with its binding partners in anti-viral responses is unknown. By performing phenotypic and functional studies of epitome-specific effectors and memory CD8 T cell populations in various situations of alternate inflammation, we propose firstly to formulate a picture of the LIGHT-HVEM-BTLA interactions that may be required to generate a virus-reactive T cell population, and secondly to formulate a rationale strategy for targeting these molecules to promote long-term protective T cell memory against respiratory viral infections. Collectively, these studies will provide new insight into the development of anti-viral CD8 T cell responses and facilitate the translational research development of novel adjuvant and vaccine strategies. PUBLIC HEALTH RELEVANCE: Many viruses utilize the respiratory tract as an entry point into the host. Respiratory viral infections are the major cause of morbidity and mortality throughout the world. Understanding how protective anti-viral CD8 T cell immunity is regulated is essential for designing safer and more effective T cell vaccines to combat respiratory pathogens and in the management of adverse immune reactions associated with immune pathology.